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Fig. 1 | Journal of Experimental & Clinical Cancer Research

Fig. 1

From: A cancer-unique glycan: de-N-acetyl polysialic acid (dPSA) linked to cell surface nucleolin depends on re-expression of the fetal polysialyltransferase ST8SIA2 gene

Fig. 1

Anti-dPSA and anti-nucleolin mAbs co-immunoprecipitates nucleolin and dPSA from the SK-MEL-28 and CHP-134 membrane fractions. Differential detergent extraction was used to separate human melanoma SK-MEL-28 cells into subcellular fractions for co-immunoprecipitation with a control murine IgG3 mAb (14C7) or anti-dPSA mAb SEAM 2 (both are subtype IgG3) linked to magnetic beads. Fractions F1, F2, and F3 are cytoplasmic, cell membrane, and nuclear fractions, respectively. Also, anti-nucleolin mAb MS-3 linked to agarose beads was used to co-immunoprecipitate proteins from the membrane fraction prepared from CHP-134 and SK-MEL-28 cells. Sections of the SimplyBlue® Coomassie-stained gels in lanes labeled F2 indicated by arrows and labeled P1–P4 of both gels were excised and proteins contained in them identified by in-gel trypsin digestion and LC/MS/MS peptide mass fingerprinting. A Proteins co-immunoprecipitated by the irrelevant IgG3 mAb. B Proteins co-immunoprecipitated by SEAM 2. C SDS-PAGE gel of proteins co-immunoprecipitated from the membrane fraction of CHP-134 (lane 1) and SK-MEL-28 (lane 2) with MS-3. (D) Western blots of MS-3-co-immunoprecipitated proteins shown in (C) stained with anti-dPSA mAb SEAM 3 (lanes 1 and 2), MS-3 (lanes 3 and 4), and irrelevant mouse IgG (lanes 5 and 6)

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